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Insights into the Mechanistic Basis of Plasmid-Mediated Colistin Resistance from Crystal Structures of the Catalytic Domain of MCR-1

DOI: 10.1038/srep39392 DOI Help

Authors: Philip Hinchliffe (University of Bristol) , Qiu E. Yang (Cardiff University) , Edward Portal (Cardiff University) , Tom Young (University of Bristol) , Hui Li (China Agricultural University) , Catherine L. Tooke (University of Bristol) , Maria J. Carvalho (Cardiff University) , Neil G. Paterson (Diamond Light Source) , Jurgen Brem (University of Oxford) , Pannika R. Niumsup (Naresuan University) , Uttapoln Tansawai (Naresuan University) , Lei Lei (China Agricultural University) , Mei Li (China Agricultural University) , Zhangqi Shen (China Agricultural University) , Yang Wang (China Agricultural University) , Christopher J. Schofield (University of Oxford) , Adrian J Mulholland (University of Bristol) , Jianzhong Shen (China Agricultural University) , Natalie Fey (University of Bristol) , Timothy R. Walsh (Cardiff University) , James Spencer (University of Bristol)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Scientific Reports , VOL 7

State: Published (Approved)
Published: January 2017
Diamond Proposal Number(s): 12342

Open Access Open Access

Abstract: The polymixin colistin is a “last line” antibiotic against extensively-resistant Gram-negative bacteria. Recently, the mcr-1 gene was identified as a plasmid-mediated resistance mechanism in human and animal Enterobacteriaceae, with a wide geographical distribution and many producer strains resistant to multiple other antibiotics. mcr-1 encodes a membrane-bound enzyme catalysing phosphoethanolamine transfer onto bacterial lipid A. Here we present crystal structures revealing the MCR-1 periplasmic, catalytic domain to be a zinc metalloprotein with an alkaline phosphatase/sulphatase fold containing three disulphide bonds. One structure captures a phosphorylated form representing the first intermediate in the transfer reaction. Mutation of residues implicated in zinc or phosphoethanolamine binding, or catalytic activity, restores colistin susceptibility of recombinant E. coli. Zinc deprivation reduces colistin MICs in MCR-1-producing laboratory, environmental, animal and human E. coli. Conversely, over-expression of the disulphide isomerase DsbA increases the colistin MIC of laboratory E. coli. Preliminary density functional theory calculations on cluster models suggest a single zinc ion may be sufficient to support phosphoethanolamine transfer. These data demonstrate the importance of zinc and disulphide bonds to MCR-1 activity, suggest that assays under zinc-limiting conditions represent a route to phenotypic identification of MCR-1 producing E. coli, and identify key features of the likely catalytic mechanism.

Journal Keywords: Antimicrobial resistance; Bacterial structural biology

Subject Areas: Medicine, Biology and Bio-materials, Chemistry

Instruments: I02-Macromolecular Crystallography , I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength)